Fc receptor function on sheep alveolar macrophages

We have examined the binding to sheep alveolar macrophages (AM) and peripheral blood polymorphonuclear leukocytes (PMN) of sheep immunoglobulin G subclasses or rabbit IgG immune complexes formed between rabbit anti-DNP IgG and DNP-bovine serum albumin. Binding studies using 125I-rabbit IgG immune complexes demonstrated 6.6 +/- 3.5 X 10(4) receptors per alveolar macrophage; these receptors bound immune complexes with an average association constant of 3.3 X 10(7) M-1. Saturation binding was achieved by 90 minutes at 4 degrees C with 6 X 10(-8) M IgG. Binding of subclasses of sheep IgG was examined by immunofluorescence. Only 10% of alveolar macrophages bound monomeric IgG1 and no binding of sheep IgG2 monomer could be demonstrated. In contrast, most peripheral blood PMN (93.0 +/- 9.5%) bound IgG2, but not IgG1. No binding to adult peripheral blood PMN of rabbit IgG immune complexes could be demonstrated. To study further the development of pulmonary host defense, we examined the expression of receptors for IgG immune complexes (Fc gamma R) on alveolar macrophages obtained from animals aged 8 through 180 days. At 8 and 21 days of age, the number of Fc gamma R varied considerably (75,000-192,000 sites per cell) and equalled or even exceeded that of adult sheep. Fc gamma R number declined by 42 and 90 days of age, where a nadir was reached (37,000 +/- 6,000 and 25,000 +/- 6,000 sites, respectively). By 180 days of age, the number of receptors had approached those of normal adult sheep (70,000 +/- 20,000 sites per cell). These studies parallel previous observations that revealed age-related differences in the phagocytic capacity of ovine alveolar macrophages.     

Human IgG1 and its Fc fragment bind with different affinities to the Fc receptors on the human U937, HL-60 and ML-1 cell lines

Measurements were made of the binding of human monomeric 125I-IgG1 and 125I-Fc to U937 cells at room temp. Analyses of the binding data showed that these cells possessed a single class of receptor (FcR) for Fc or IgG and, although both ligands were found to bind to the same number of sites per cell, Fc was found to bind with about twice the affinity of IgG. At 20 degrees C estimates of the forward rate constants and the dissociation rate constants for IgG and Fe were 1.13 and 3.65 X 10(7) M-1 min-1 and 0.33 and 0.57 X 10(-2) min-1 respectively. Independent determinations of the association constants (Ka) under the same experimental conditions gave values of 0.98 X 10(9) M-1 for IgG and 3.1 X 10(9) M-1 for Fc. Thus the Fc fragment of IgG appears to bind to U937 FcR at 3-4 times the rate of IgG and to dissociate at about twice the rate, resulting in higher values of Ka for the Fc-FcR than for the IgG-FcR interaction. Also, in competitive-binding experiments and in EA rosette inhibition assay the Fc fragment was consistently found to be more efficient in FcR binding than IgG. Similar results were obtained using HL-60 and ML-1 cells which possess FcR like those on U937 cells and with IgG1 and Fc prepared from other myelomas. IgG and Fe which had undergone mild reduction and alkylation bound to the same number of FcR per U937 cell as the non-reduced ligands but the affinity of binding was diminished to a similar degree with both ligands, suggesting that the major effect of cleavage of the interchain disulfide bonds on cytophilic binding is due to alteration of the native quaternary relationships of the C gamma 2 and C gamma 3 domains.     

Fc gamma receptor recognition of IgG ligand by human monocytes and macrophages

We examined the binding characteristics of human monocytes and macrophages with the IgG ligands, human monomeric IgG and a small human IgG aggregate, trimeric IgG. Our purpose was to utilize fresh monocytes, in vitro cultured monocytes, and alveolar macrophages in direct and indirect binding experiments. Freshly isolated monocytes expressed only a single binding site for IgG monomer and IgG trimer. In contrast, in vitro cultured monocytes, gamma-interferon-treated monocytes, and freshly isolated alveolar macrophages expressed a single binding site for IgG monomer and, in addition, a high and low affinity binding site for IgG trimer. The high affinity binding site for IgG trimer (Kd approximately equal to 1 nM) appeared identical to the binding site for IgG monomer. The low affinity binding site for IgG trimer (Kd = 50 to 250 nM) appeared to be due to Fc gamma RII, because antibody to Fc gamma RII inhibited its expression. Since Fc gamma RII, in contrast to Fc gamma RI, does not bind monomeric IgG, the data suggest that this low affinity receptor for trimeric IgG, Fc gamma RII, can bind low molecular weight circulating immune complexes at concentrations 10- to 100-fold lower than Fc gamma RI. Thus, these studies suggest that at 37 degrees C, macrophage Fc gamma RII may play a functional role in the recognition of small molecular weight immune complexes.     

Binding kinetics of monomeric and aggregated IgG to Kupffer cells and hepatocytes of mice

The binding kinetics of human monomeric IgG and stable heat-aggregated IgG (A-IgG) to Fc receptors of hepatocytes and Kupffer cells isolated from mice was studied. After injection of radiolabelled proteins the 60-70% of hepatic uptake was recovered in parenchymal cells (hepatocytes). In experiments in vitro the A-IgG bound in larger amounts to hepatocytes and Kupffer cells than monomeric IgG. The association rate constants of aggregates were somewhat higher for Kupffer cells than for hepatocytes whereas the percentage uptake of aggregates by Kupffer cells was only 5-15% of that of hepatocytes. The equilibrium constants of aggregates binding to both cells amounted to 0.4-1 X 10(8) M-1 for A-IgG compared with an equilibrium constant for monomeric IgG of 1-2 X 10(7)M-1. The maximum number of IgG and A-IgG molecules bound per cell was higher on hepatocytes (mean 14 X 10(6)) than on Kupffer cells (mean 2 X 10(5)) which is in agreement with the higher binding capacity of hepatocytes for these proteins observed in vivo and in vitro experiments. The ability to compete for receptor binding seemed to reside exclusively in the Fc portion of IgG since F(ab')2 fragments of IgG failed to inhibit labelled monomeric IgG or A-IgG. The receptor seems to be specific for IgG since unlabelled monomeric IgA demonstrated no binding inhibition of labelled IgG or A-IgG on hepatocytes and Kupffer cells. The overall results further suggest that hepatocytes might through Fc receptors play a collaborative role with the mononuclear phagocytic system in the clearance of circulating immune complexes.     

The binding of rabbit IgG and its enzymatically derived fragments to homologous peritoneal macrophages.

Rabbit IgG and its Fab, Fc and pFc' fragments, prepared by papain or peptic digestion, were assayed for binding to homologous peritoneal macrophages. The binding affinity of IgG for the peritoneal macrophages (Ka = 5.9 +/- 1.6 x 10(5) L/M) was comparable to that recorded with alveolar macrophages (7.6 +/- 1.8 x 10(5) L/M, Arend & Mannik, 1973) but the number of receptor sites per peritoneal cell (4.6 +/- 2.1 x10(6)) was about four-fold greater than on the latter. Of the fragments, only Fc bound to macrophages with an affinity comparable to intact IgG; pFc' bound weakly and Fab was totally inactive. These data, taken with a recent study involving rabbit IgG and guinea-pig macrophages (Ovary, Saluk, Quijada & Lamm, 1976), indicate that the primary IgG binding site for macrophages is located in the C gamma 2 domain.     

The spontaneous ability of normal human IgG to inhibit the Fc receptors of normal human monocytes is related to their binding capacity to lectins

The lectin-binding capacity of 96 normal human IgG, assessed by solid-phase radioimmunoassay, strikingly varied according to the lectin considered. Indeed, half of the IgG samples exhibited peanut agglutinin (PNA)- and pokeweed mitogen-specific binding capacities superior or equal to 4%, whereas less than 15% of IgG similarly bound to concanavalin A (Con A) and to phytohemagglutinin. The ability of those IgG to inhibit the Fc receptor (Fc-R) function of human monocytes, measured by a classical rosette assay, was inversely correlated to their binding ratios to PNA and Con A only. By affinity chromatography, three groups of IgG were separated: the IgG purified on agarose-PNA columns slightly reduced the Fc-R function (40-45% inhibition); the IgG purified on Sepharose-Con A columns exhibited the highest inhibitory properties (80-85% inhibition); the IgG that did not bind to PNA and Con A columns possessed intermediate inhibitory properties (65-70% inhibition). The different effect of IgG on Fc receptors was conserved when monocytes were first treated by trypsin and was unrelated to their specific binding to human monocytes, to their subclasses, and to their C1q- or protein A-binding capacities. Incubation of monocytes with D-galactose (10 mM) significantly improved their capacity to form IgG rosettes, whereas their incubation with D-mannose (10 mM) significantly reduced the Fc-R function. Scatchard plots of 125I-IgG1 myeloma protein binding to monocytes were linear under basal conditions, as well as after a prior incubation of the cells with D-galactose or D-mannose. Monocytes bound about 16,000 molecules of IgG1 per cell in each instance. In contrast, the mean association constant (Ka) for IgG1 binding was 2.59 +/- 0.50 X 10(8) M-1 under basal conditions, 4.4 +/- 0.75 X 10(8) M-1 after D-galactose incubation, and 1.35 +/- 0.50 X 10(8) M-1 after D-mannose incubation. These data suggest that the level of human monocyte Fc-R function blockade induced by human IgG depends mainly on the presence of "accessible" galactosyl or mannosyl residues in the Fc domain and that the modulation of the Fc-R function induced by these carbohydrates is due to a change in the affinity rather than in the number of single class of high-affinity binding sites.     

Possible errors in the calculation of cell-binding parameters exemplified by the analysis of the IgG-Fc receptor system

The binding parameters of monomeric and polymeric (immune complex with a molecular weight of 500,000 daltons) rabbit IgG to homologous Fc receptor-bearing alveolar macrophages were estimated, using corrected values for IgG and cell concentrations. Considering the maximum percentage of monomeric IgG binding to cells (2.7%) and the maximum percentage of cells binding monomeric IgG (32%) instead of the IgG and cell concentrations in the initial reaction mixture, a 36-fold increase of the equilibrium constant (K) (from 0.6 x 10(6) L/M to 21.3 x 10(6) L/M) and a 3-fold increase of the maximum number of IgG molecules able to bind to a single cell (n) (from 7.8 x 10(5) to 23.7 x 10(5] were registered. Since more than 60% of the polymeric IgG is bound to 46% of the macrophage population, the values of K (from 10.8 x 10(6) L/M to 15.6 x 10(6) L/M) and n (from 4.3 x 10(5) to 9.4 x 10(5] are only doubled by using the corrected values for IgG and cell concentrations. It results that the cytophilic fraction of the monomeric IgG, representing only 2.7% of total IgG, has a slightly higher affinity for the Fc receptors than the polymeric IgG. By considering the actual number of macrophages which bind IgG it is evident that the number of Fc receptors per cell is higher than that determined by the usual procedure which does not take into account cellular heterogeneity.     

Isolation of a 30 kDa immunoglobulin binding protein from Pseudomonas maltophilia

We have demonstrated that Pseudomonas maltophilia (ATCC No. 13637) possesses an exposed, immunologically accessible protein which binds to the Fc region of several species of immunoglobulins. Whole bacteria suspensions were incubated for 18 h with purified 125I-labelled antibodies with and without added non-labelled immunoglobulins. The suspensions were centrifuged for 30 min and the pellet containing bacteria was assessed for radioactivity. Using this crude assay, the whole organism bound 125I-labelled rabbit and mouse immunoglobulins and the purified Fc portion of human IgG. All of these labelled preparations were competitively displaced by unlabelled rabbit and mouse immunoglobulins, and Fc of human IgG, as well as human immunoglobulin subclasses. The organism was sonicated to solubilize this immunoglobulin binding protein. Using this sonicated preparation, it was shown that unlabelled Fc of IgG, unlabelled mouse and rabbit immunoglobulins, all competitively displaced 125I-labelled human Fc of IgG in a dose-response manner. A partially purified protein was prepared by Sephacryl S-300 followed by Sephadex G-100 column chromatography. This preparation was incubated with 125I-Fc gamma and with the following purified unlabelled preparations: F(ab')2 of IgG, Fc of IgG, murine monoclonal IgA, IgG1, IgG2, IgG3, and IgG4. All except F(ab')2 of IgG produced dose response competitive displacement. The molecular weight, as estimated by SDS-PAGE and Western blot, was 30,000 daltons. In Western blots, Fc gamma, murine monoclonal IgA, and human immunoglobulin subclasses, all showed affinity for the immobilized protein. Human F(ab')2 fragments did not show affinity for the protein. Radioiodinated pseudomonal Ig-binding protein showed affinity for human IgG coupled to Sepharose, and was displaced by unlabelled pseudomonal Ig-binding protein. Scatchard analysis of binding showed two binding affinities: two distinct types of Ig-binding proteins were obtained, a high affinity with Kd = 1.54 x 10(-10) and a lower affinity with Kd = 2.36 x 10(-8). This immunoglobulin binding protein may be useful in immunoglobulin purification or identification.     

Isolation of a rabbit IgG fraction with cytophilic properties

About 15% of rabbit IgG loaded on a column of Con A-Sepharose 4B was found to be specifically bound to the column due to a structural variation in its carbohydrate moiety. The Con A-retained rabbit IgG contained a higher amount of neutral hexoses than the initial IgG but its molecular weight, antigenic structure and half-life were identical or similar. The Con A-retained rabbit IgG has an affinity for the Fc receptor-bearing homologous macrophages which is 10 times higher than that of the initial IgG. The IgG fraction not retained on Con A-Sepharose is practically devoid of binding ability. These results suggest that the Con A-bound IgG may represent the cytophilic fraction of monomeric IgG responsible for the binding of IgG to Fc receptor-bearing cells.     

Comparison of the Binding of Radiolabelled Human IgG and Fc Fragments to Murine Spleen Cells

The binding properties of an IgG1 human myeloma protein, normal IgG, and the Fc and Fab fragments of each were compared in cultures of murine spleen cells. Both 125I-labelled IgG and Fc fragments bound to splenic lymphocytes, whereas Fab fragments did not bind significantly at the highest concentrations tested. On a molar basis, more Fc bound than intact IgG. According to Scatchard plot analysis, the affinity constand of IgG1 was 1.5 x 10(6) +/- 1 x 10(5) L/M and that of the Fc fragments was 7.8 x 10(5) +/- 2.6 x 10(5) L/M. Approximately 25,000 binding sites/cell were calculated for IgG1 and 102,000 for Fc. Deaggregation of the Fc preparation did not change these values, suggesting that the difference in binding of IgG and Fc did not result from Fc aggregation. Unlabelled IgG inhibited about 25% of the labelled Fc binding, whereas unlabelled Fc inhibited approximately 80% of the labelled Fc binding. IgG antigen-antibody complexes, however, inhibited 75% of the Fc binding. In the reciprocal experiment both intact IgG and Fc inhibited the binding of labelled IgG by 100%. The major cell population that bound IgG and Fc fragments in the spleen cell preparation were the B lymphocytes. Removal of macrophages did not significantly affect the binding of labelled Fc fragments. In addition, T-cell-enriched populations bound an insignificant quantity of Fc fragments.     

Non-complement-dependent adsorption of soluble immune complexes to human red cells

Heat aggregated IgG and soluble immune complexes (IC) prepared by combining human serum albumin with rabbit anti-serum albumin and tetanus toxoid with rabbit antiserum to tetanus toxoid were shown to bind to human O+ RBC. The binding was greater for soluble IC prepared at antigen excess, and although it was usually maximal when IC were pre-opsonized, it could also be demonstrated using non-opsonized heat aggregated IgG or soluble IC prepared in the absence of complement. These observations suggest that two types of receptors may be involved in binding of soluble IC to human RBC: the classical C3b receptor, and a non-complement-dependent receptor, perhaps recognizing the Fc region of the immunoglobulin molecule exposed after heat aggregation or antigen-antibody reaction.     

Species specificity in the binding of IgG to macrophages.

The binding of human IgG1 and IgG3 and rabbit IgG to guinea-pig peritoneal macrophages was examined, and differences between the species, in terms of their binding mechanisms, were characterized. Rabbit IgG bound with high affinity (Kass = 3.11 +/- 0.45 x 10(6) M-1) to a finite number of receptor sites per cell (1.26 +/- 0.29 x 10(6)) and competitively inhibited the binding of guinea-pig IgG2. Heterogeneity in binding, with distinct high and low affinity components, was observed when human IgG3 was reacted with guinea-pig macrophages, while human IgG1 exhibited only low affinity binding. Neither human IgG subclass competed effectively with guinea-pig IgG2 for its cell receptor. Thus, rabbit IgG appeared to cross-react with a macrophage receptor for guinea-pig immunoglobulin, whereas the human IgG subclasses bound to macrophage membrane components that remained undefined.     

Human monocyte binding of homologous monomer and complexed IgG.

Human peripheral blood monocyte binding of homologous IgG has been studied using monomer and covalently cross-linked dimer and trimer. A competitive radioassay using enriched monocytes was employed to determine the association constant (Ka) and the number of receptor sites per cell. Monomer and dimer bound with very similar Ka values (7.15 X 10(7) and 7.14 X 10(7)LM-1) whereas a slightly higher figure was obtained with trimer (8.9 X 10(7)LM-1). The number of receptor sites for dimer and trimer (90.5 and 107.5 X 10(3)/cell, respectively) were much greater than for monomer (14.5 X 10(3)/cell). Inhibition of dimer and trimer binding by monomer gave heterogenous Scatchard plots, each with two components. These findings suggest that human peripheral blood monocytes possess two types of Fc receptor, one of high affinity binding monomer and complexes and one of low affinity which predominantly binds complexes.     

The human monocyte-like cell line THP-1 expresses Fc gamma RI and Fc gamma RII.

THP-1 cells are a monocyte-like cell line derived from a patient with acute monocytic leukemia and unlike other leukemic cell lines has a normal diploid karyotype. We have characterized Fc gamma R expression on this cell line by flow cytometry, radiolabeled IgG1 and monoclonal antibody (mAb) binding assays, and biochemical analysis. Flow cytometric analysis of THP-1 cells with anti-Fc gamma RI, II, and III mAb, and a rabbit anti-Fc gamma RIII F(ab')2 demonstrated that only Fc gamma RI and Fc gamma RII are expressed by these cells. A panel of anti-Fc gamma RIII mAb (anti-CD16) failed to bind to THP-1 cells. Biochemical studies identified polypeptides of 64 to 78 kDa (Fc gamma RI) and of 42 to 53 kDa (Fc gamma RII). Fc gamma R expression was determined by binding of radioiodinated human IgG1 (to detect Fc gamma RI), mAb IV.3 (to detect Fc gamma RII), or rabbit IgG immune complexes. Thirty-five thousand high affinity binding sites (dissociation constant [KD] = 4.22 x 10(-9) M) for IgG1 were found on THP-1 cells. Interferon-gamma (IFN gamma) upregulated Fc gamma RI expression by THP-1 cells 2.8-fold, whereas Fc gamma RI on U937 cells was increased six- to eight-fold by this cytokine. Phorbol myristate acetate (PMA), tumor necrosis factor-alpha (TNF alpha), and vitamin D3 had no effect on IgG1 binding by THP-1 cells. Fifty thousand IgG molecules in immune complexes bound to THP-1 cells. IFN gamma treatment increased this binding by four-fold, PMA treatment resulted in a 50% increase in the number of IgG immune complexes bound, whereas vitamin D3 treated THP-1 cells bound half as many IgG immune complexes as control cells. Binding assays utilizing mAb IV.3 identified 50,000 sites per cell. Treatment of THP-1 cells with IFN gamma, TNF alpha, PMA, or vitamin D3 had no effect on Fc gamma RII expression. That Fc gamma RI plays a predominant role in immune complex binding was demonstrated by inhibition studies. Human IgG1 as well as mouse IgG2a mAb to Fc gamma RII inhibited immune complex binding by 76 to 84%, whereas mouse IgG1 mAb to Fc gamma RII had minimal effect on immune complex binding. Fc gamma R expression may not be linked to differentiation of THP-1 cells since only 1,25 vitamin D3 was able to induce the expression of CD14, a marker of mature monocytic phenotype.     

The human mononuclear phagocyte high-affinity Fc receptor, FcRI, defined by a monoclonal antibody, 10.1

The properties of the mononuclear phagocyte (Mph) high-affinity Fc receptor, FcRI, were investigated using a novel monoclonal antibody (mAb) designated 10.1. This receptor was shown to be a protein of 71 kDa, presented chiefly on monocytes and the myeloid cell lines U937 and HL60. mAb 10.1 inhibited the binding to Mph of erythrocytes opsonized with rabbit IgG or human IgG3. It also blocked T cell proliferation induced by murine CD3 mAb of the IgG2a but not the IgG1 subclass. These results suggest that rabbit IgG, human IgG3 and murine IgG2a all bind to FcRI in a similar manner and that mAb 10.1 reacts with an epitope on FcRI near to the binding site for the Fc region of IgG. In addition, although it is well known that FcRI has a high affinity for both monomeric human IgG1 and IgG3, we show in this study that while erythrocytes opsonized with human IgG3 bind to Mph, equivalent cells opsonized with IgG1 surprisingly do not. These results define further the nature of the constraints on the interaction between Mph FcRI and particular IgGs.     

Histidine-rich glycoprotein regulates the binding of monomeric IgG and immune complexes to monocytes.

Histidine-rich glycoprotein (HRG) is a relatively abundant plasma protein which we have shown previously inhibits the formation of insoluble immune complexes (IC). In this study we examined the ability of HRG to regulate the binding of monomeric IgG and IC to monocytes. Initial studies demonstrated that HRG interacts with FcgammaRI on the monocytic cell line THP1 and blocks the binding of monomeric IgG to these cells. However, despite totally blocking the binding of monomeric IgG to FcgammaRI, pre-incubation of THP1 cells with HRG had no effect on the binding of IC to these cells. In contrast, depending on the HRG:IgG molar ratio, pre-incubation of monomeric IgG with HRG resulted in either enhanced or reduced IgG binding to FcgammaRI. Similarly, under certain highly defined conditions, incorporation of HRG in IgG-containing IC potentiated the binding of IC to THP1 cells. The key conditions involved incorporating approximately equimolar concentrations of HRG and IgG in the IC, the IC being formed at a near equivalence antigen:antibody ratio and usually physiological concentration (20 microM) of Zn(2+) being present. Collectively these observations indicate that HRG is an important regulator of IC uptake by monocytes. Thus HRG can interact with FcgammaRI on monocytes and block monomeric IgG binding, whereas when incorporated in IgG containing IC, HRG can enhance the uptake of IC by monocytes, probably via its heparan sulfate binding domain.     

Highly reduced binding to high and low affinity mouse Fc gamma receptors by L234A/L235A and N297A Fc mutations engineered into mouse IgG2a

The effects of the Fc silencing mutations such as leucine (L) to alanine (A) substitution at the position 234 and 235 (LALA) and the alanine (A) to asparagine (N) substitution at position 297 (N297A) are well investigated for human IgG. However, the effects of the same two silencing Fc mutations in a mouse IgG backbone are not yet well investigated in respect to binding to mouse Fc gamma receptors (FcγRs), complement and subsequent effector functions. By using a mouse IgG2a tool antibody directed against mouse OX40L, we demonstrate a strongly reduced binding of the two Fc mutants to high and low affinity recombinant and cell expressed mouse FcγRs, when compared to the mouse IgG2a with the wild type (wt) backbone. Reduced FcγR binding by the two investigated Fc mutants could further be confirmed on primary mouse macrophages expressing their native FcγRs. In addition, we reveal that the LALA and N297A mutations in the mIgG2a also slightly reduced binding to C1q of human origin. Thus, here we provide experimental evidence that the two investigated Fc mutations in the mouse IgG backbone lead to similar “silencing” properties as previously demonstrated for the human IgG and thus represent a useful method to alter effector functions in tool antibodies to be used in mouse models.     

A kinetic study of human IgG binding to placental Fc gamma-receptor

Various methods for studying the IgG interaction with its placental receptor have been examined to establish kinetic parameters. IgG binding has an average rate constant k1 of (2.57 +/- 0.94) x 10(7)/M per min. However, high displacement doses (1000 micrograms) of unlabelled IgG resulted in a curvilinear dissociation curve with two binding sites: one with a fast dissociation rate constant, k2 of 0.100/min and a slow one with k2 of 0.010/min. IgG binding was associated with a high average affinity of (3.70 +/- 1.65) x 10(8)/M and a total number of receptor binding sites of (2.07 +/- 0.93) x 10(14) sites/mg of membrane protein, in close agreement with previous results. The stoichiometry of IgG to Fc gamma-receptor was established as a 4:1 binding ratio from log dose-response curves as opposed to a 1:1 binding ratio from previous reports.     

IgG Binding Sites on Human FCγ Receptors

The structure for the three human Fcγ receptors classes FCγRI (CD64), FCγRII (CD32) and FCγRIII (CD16) has been well characterized. Here the IgG binding sites on FCγRII and FCγRIII with their responsive FG, BC and C'/E loops on the membrane proximal domains are described in detail. For FCγRI the second extracellular domain is suggested as a key structure of IgG binding. The lower hinge regions of human and murine IgG binding to these Fc receptors and their structural relationship in FcγR-IgG interactions are discussed. The potential of inhibiting the pathophysiological effects of Fcγ receptors by blocking studies are considered for future therapeutic modalities.     

Functional affinity constants of subfragments of immunoglobulin G for Clq

The functional affinity constants for Clq of subfragments if IgG1 representing the C gamma 2 (c gamma 2III) region or the whole C gamma 3 region of Fc (pFc'), have been measured by examining the ability of these fragments to inhibit the interaction between radiolabelled Clq and glutaraldehyde-treated human erythrocytes or aggregated human IgG. The value of the functional affinity constant for the C gamma 2III fragment was the same as that for Fc and that determined previously for monomeric IgG, indicating that all the elements necessary for Clq binding are contained in a single C gamma 2 domain. The pFc' fragment was inactive but a more degraded trypsin fragment from this region, at C gamma 3, showed the same affinity of binding for Clq as the C gamma 2III and Fc. These results confirm earlier findings that it is not combination of residues in the C gamma 2 which bind Clq which is responsible for their activity but their accessibility.